Linked ligation
AssigneeNcan Genomics, Inc.
InventorsAndrea Marziali, Joel Pel
Life of this patent
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Abstract
The invention generally relates to capturing, amplifying, and sequencing nucleic acids. In certain embodiments, copies of the sense and antisense strands of a duplex template nucleic acid are captured using linked capture probes and multiple binding and extension steps to improve specificity over traditional single binding target capture techniques. Methods of seeding sequencing clusters with sense and antisense strands of a target nucleic acid are also disclosed including identifying the strands using sense-specific barcodes and confirming base calls using two sense-specific sequencing reads. Linked adapters may be used to increase adapter ligation selectively or efficiency and yield.
Claims21
A method for of selectively ligating adapters to a target nucleic acid, the method comprising:
- providing a first linked ligation adapter comprising a first adapter linked to a first probe complementary to a first portion that is internal to 5′ and 3′ ends of the target nucleic acid, wherein the first probe is linked to the first adapter by a first linker;
- hybridizing the first probe to the target nucleic acid;
- and
- ligating the first adapter to a first terminus of the target nucleic acid to produce a first ligation product.
A method of enrichment of a target nucleic acid, the method comprising:
- (a) producing a first mixture of ligation products by combining a mixture comprising nucleic acid fragments and a first linked ligation adapter comprising a first adapter linked to a first probe complementary to a first portion internal to 5′ and 3′ ends of the target nucleic acid, wherein the first adapter is linked to the first probe by a first linker;
- (b) producing a second mixture of ligation products by combining a mixture comprising the first mixture and a second linked ligation adapter comprising a second adapter linked to a second probe complementary to a second portion of the target nucleic acid by a second linker;
- and
- (c) removing fragments from the second mixture that are not ligated to the first or the second linked ligation adapters.
Dependencies are inferred from the claim text.